Key Laboratory of Natural Medicines of the Changbai Mountain, Ministry of Education, Molecular Medicine Research Center, College of Pharmacy, Yanbian University, Yanji 133002, Jilin, China.
Key Laboratory of Natural Medicines of the Changbai Mountain, Ministry of Education, Molecular Medicine Research Center, College of Pharmacy, Yanbian University, Yanji 133002, Jilin, China.
Int Immunopharmacol. 2022 Nov;112:109176. doi: 10.1016/j.intimp.2022.109176. Epub 2022 Sep 5.
Toxoplasma gondii (T. gondii) is a neurotropic obligate intracellular parasite that can activate microglial and promote neuronal apoptosis, leading to central nervous system diseases. The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome signaling complex plays a key role in inducing neuroinflammation. Our previous studies have found that ginsenoside Rh2 (GRh2) inhibits T. gondii infection-induced microglial activation and neuroinflammation by downregulating the Toll-like receptor 4/nuclear factor-kappa B signaling pathway. However, whether GRh2 reduces T. gondii infection-induced neuronal injury through actions on microglial NLRP3 inflammasome signaling has not yet been clarified.
In this study, we employed T. gondii RH strain to establish in vitro and in vivo infection models in BV2 microglia cell line and BALB/c mice. Molecular docking, localized surface plasmon resonance assay, quantitative competitive-PCR, ELISA, western blotting, flow cytometric analysis, and immunofluorescence were performed.
Our results showed that GRh2 alleviated neuropathological damage and neuronal apoptosis in cortical tissue of T. gondii-infected mice. GRh2 and CY-09 (an inhibitor of NLRP3) exhibited potent anti-T. gondii effects through binding T. gondii calcium-dependent protein kinase 1 (TgCDPK1). GRh2 decreased Iba-1 (a specific microglial marker) and NLRP3 inflammasome signaling pathway-related protein expression by binding NLRP3. Co-culture of microglia/primary cortical neurons revealed that T. gondii-induced microglial activation caused neuronal apoptosis, but GRh2 reduced this effect, consistent with the effects of CY-09.
Taken together, our results show that GRh2 has a protective effect against T. gondii infection-induced neuronal injury by binding TgCDPK1 and NLRP3 to inhibit NLRP3 inflammasome signaling pathway in microglia.
刚地弓形虫（T. gondii）是一种神经嗜性的必需的细胞内寄生虫，它可以激活小胶质细胞并促进神经元凋亡，导致中枢神经系统疾病。NOD 样受体家族含 pyrin 域蛋白 3（NLRP3）炎性小体信号复合物在诱导神经炎症中起着关键作用。我们之前的研究发现，人参皂苷 Rh2（GRh2）通过下调 Toll 样受体 4/核因子-κB 信号通路抑制弓形虫感染诱导的小胶质细胞激活和神经炎症。然而，GRh2 是否通过作用于小胶质细胞 NLRP3 炎性小体信号来减轻弓形虫感染诱导的神经元损伤尚未阐明。
本研究采用弓形虫 RH 株在 BV2 小胶质细胞系和 BALB/c 小鼠中建立体外和体内感染模型。进行分子对接、定位表面等离子体共振分析、定量竞争-PCR、ELISA、western blot、流式细胞术分析和免疫荧光。
我们的结果表明，GRh2 减轻了弓形虫感染小鼠皮质组织的神经病理损伤和神经元凋亡。GRh2 和 CY-09（NLRP3 的抑制剂）通过与弓形虫钙依赖性蛋白激酶 1（TgCDPK1）结合表现出强大的抗弓形虫作用。GRh2 通过与 NLRP3 结合降低 Iba-1（一种特异性小胶质细胞标志物）和 NLRP3 炎性小体信号通路相关蛋白的表达。小胶质细胞/原代皮质神经元共培养显示，弓形虫诱导的小胶质细胞激活导致神经元凋亡，但 GRh2 降低了这种作用，与 CY-09 的作用一致。
综上所述，我们的研究结果表明，GRh2 通过与 TgCDPK1 和 NLRP3 结合抑制小胶质细胞中 NLRP3 炎性小体信号通路，对弓形虫感染诱导的神经元损伤具有保护作用。
